Ekologické zemědělství - informační bulletin č.11: Přímý prodej

Jednotným tématem informačního bulletinu č. 11 je Přímý prodej biopotravin: Předpoklady pro přímý prodej. Přímý prodej produktů rostlinného původu. Jakost ovoce z ekologického pěstování. BIO na tržnici v St.Gallenu. Přímý prodej produktů živočišného původu. Marketing v přímém prodeji. Jednotlivé způsoby přímého prodeje ekologických produktů. Příklady ekologicky hospodařících zemědělců s přímým prodejem. Společné ztržňování bioproduktů

Modelling of orbital deformation using finite-element analysis

The purpose of this study was to develop a three-dimensional finite-element model (FEM) of the human orbit, containing the globe, to predict orbital deformation in subjects following a blunt injury. This study investigated the hypothesis that such deformation could be modelled using finite-element techniques. One patient who had CT-scan examination to the maxillofacial skeleton including the orbits, as part of her treatment, was selected for this study. A FEM of one of the orbits containing the globe was constructed, based on CT-scan images. Simulations were performed with a computer using the finite-element software NISA (EMRC, Troy, USA). The orbit was subjected to a blunt injury of a 0.5 kg missile with 30 m s(−1) velocity. The FEM was then used to predict principal and shear stresses or strains at each node position. Two types of orbital deformation were predicted during different impact simulations: (i) horizontal distortion and (ii) rotational distortion. Stress values ranged from 213.4 to 363.3 MPa for the maximum principal stress, from −327.8 to −653.1 MPa for the minimum principal stress, and from 212.3 to 444.3 MPa for the maximum shear stress. This is the first finite-element study, which demonstrates different and concurrent patterns of orbital deformation in a subject following a blunt injury. Finite element modelling is a powerful and invaluable tool to study the multifaceted phenomenon of orbital deformation

Grundlagen für die Düngung der Obstkulturen - Kernobst, Steinobst, Kiwi, Strauchbeeren

Die Anbausysteme im modernen Erwerbsobstbau haben sich verändert in den letzten Jahren. Das ist der Hauptgrund für eine komplett überarbeitete Neuausgabe der bisherigen Richtlinien (Heller et al. 1993; Commission romande de fumure, sous-commission arboricole 1993; Ryser et al. 1995). Der Wandel von Sorten, Veredlungsunterlagen, Anbauformen und Pflanzsystemen beeinflusst die Gesamtmenge an Nährstoffen, welche eine moderne Obstanlage braucht, die Aufteilung der Nährstoffe auf die verschiedenen Organe der Pflanze und auch die Nährstoffmenge, welche mit dem Ernteprodukt die Obstanlage verlässt. Damit stellt sich die Frage, wie moderne Anlagen bedarfsgerecht ernährt werden können. Die Anbausysteme sind dynamischer geworden, d.h. sie wandeln sich rascher als früher durch die dauernde Anpassung der Erwerbsanbaupraxis an aktuelle Marktbedürfnisse, technischen Fortschritt und Standortverhältnisse. Ziel moderner Obstproduzentinnen und -produzenten ist die Erzeugung eines Optimums an Fruchtqualität auf hohem Ertragsniveau unter Vermeidung der Alternanz. Die Baumernährung moderner Obstanlagen muss flexibel an die gegebenen Standortverhältnisse und Anbauziele angepasst sein. Die Gesellschaft verlangt zudem, dass mit den Ressourcen im Anbausystem schonend umgegangen wird

Chromosome Missegregation Associated with RUVBL1 Deficiency

RUVBL1 (RuvB-like1) and RUVBL2 (RuvB-like 2) are integral components of multisubunit protein complexes involved in processes ranging from cellular metabolism, transcription and chromatin remodeling to DNA repair. Here, we show that although RUVBL1 and RUVBL2 are known to form heterodimeric complexes in which they stabilize each other, the subunits separate during cytokinesis. In anaphase-to-telophase transition, RUVBL1 localizes to structures of the mitotic spindle apparatus, where it partially co-localizes with polo-like kinase 1 (PLK1). The ability of PLK1 to phosphorylate RUVBL1-but not RUVBL2-in vitro and their physical association in vivo suggest that this kinase differentially regulates the function of the RuvB-like proteins during mitosis. We further show that siRNA-mediated knock-down of RuvB-like proteins causes severe defects in chromosome alignment and segregation. In addition, we show that the ATPase activity of RUVBL1 is indispensable for cell proliferation. Our data thus demonstrate that RUVBL1 is essential for efficient mitosis and proliferation

RUVBL1 depletion affects the length of mitosis and results in lagging chromosomes.

<p><b>(A)</b> HeLa cells were transfected with the indicated siRNA oligos and analyzed by RT-PCR and immunoblot 48 h post-transfection. <b>(B)</b> Confocal live imaging after siRNA-mediated knock-down was performed. The figure shows stills of control or RUVBL1 siRNA-treated cells. Lagging chromosomes are indicated with arrowheads. DNA is shown in cyan and α-tubulin in red. <b>(C)</b> Early mitotic progression was analyzed by measuring the time from prophase (nuclear envelope breakdown: NEB) to anaphase onset (left panel). Mitotic exit was estimated by measuring the period of time from anaphase onset until accomplished cytokinesis (right panel). The bottom and top of the boxes represent the first and third quartiles, respectively. Horizontal lines inside the boxes represent the median of the data points (n = 100). Whiskers span the 10th and 90th percentiles, with individual dots showing data points that lie outside of these percentiles. Statistical significance was determined by Mann–Whitney test and <i>p</i>-values are indicated above. <b>(D)</b> Occurrence of aberrant mitotic phenotypes was quantified by analyzing 100 cell divisions in each cell line.</p

<i>In vitro</i> phosphorylation of RUVBL1 by PLK1.

<p><b>(A)</b> Protein sequences were obtained from <a href="http://www.ncbi.nlm.nih.gov/" target="_blank">http://www.ncbi.nlm.nih.gov</a> and scanned for PLK1 consensus motifs [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0133576#pone.0133576.ref037" target="_blank">37</a>]. Corresponding sequences of human RUVBL2 are shown by comparison below the alignments. Crucial amino acids are shown in gray. <b>(B)</b> The indicated amounts of His-tagged RUVBL1 or RUVBL2 were incubated with PLK1 in the presence of [γ-³²P]ATP and protein phosphorylation was monitored by autoradiography. <b>(C)</b> Bands corresponding to phosphorylated RUVBL1 were excised and in-gel digested with trypsin. Eluted peptides were separated in two dimensions by their hydrophobicity and charge. Potential sequences were assigned to the obtained phosphopeptide patterns after autoradiography. Trypsin digestion sites are indicated with K•X. <b>(D)</b> Purified His-tagged RUVBL1 mutants were incubated with PLK1 in the presence of [γ-³²P]ATP and protein phosphorylation was monitored by autoradiography.</p

RUVBL1 depletion affects the length of mitosis and results in lagging chromosomes.

<p><b>(A)</b> HeLa cells were transfected with the indicated siRNA oligos and analyzed by RT-PCR and immunoblot 48 h post-transfection. <b>(B)</b> Confocal live imaging after siRNA-mediated knock-down was performed. The figure shows stills of control or RUVBL1 siRNA-treated cells. Lagging chromosomes are indicated with arrowheads. DNA is shown in cyan and α-tubulin in red. <b>(C)</b> Early mitotic progression was analyzed by measuring the time from prophase (nuclear envelope breakdown: NEB) to anaphase onset (left panel). Mitotic exit was estimated by measuring the period of time from anaphase onset until accomplished cytokinesis (right panel). The bottom and top of the boxes represent the first and third quartiles, respectively. Horizontal lines inside the boxes represent the median of the data points (n = 100). Whiskers span the 10th and 90th percentiles, with individual dots showing data points that lie outside of these percentiles. Statistical significance was determined by Mann–Whitney test and <i>p</i>-values are indicated above. <b>(D)</b> Occurrence of aberrant mitotic phenotypes was quantified by analyzing 100 cell divisions in each cell line.</p

ATPase activity of mammalian RUVBL1 is indispensable for cell proliferation.

<p><b>(A)</b> FLAG-tagged RUVBL1 was transiently expressed in 293T cells and protein extracts were prepared 48 h after transfection. Anti-FLAG beads were used to isolate the tagged RUVBL1. Silver staining PAGE shows the purity of the isolated material. <b>(B)</b> ATPase activity was measured by incubating purified FLAG-tagged RUVBL1 with [γ-³²P]ATP for the times indicated, either in the presence or absence of ssDNA. <b>(C)</b> U2OS T-REx cells carrying stably-integrated, inducible shRNA-resistant murine RuvBL1 variants, were treated with doxycycline for 96 h (lane 2 and 5). In addition, the cell lines were stably-transfected with an inducible shRNA construct that enables downregulation of endogenous RUVBL1 (lane 3 and 6) after doxcycycline addition. Parental cells are shown for comparison (lanes 1 and 4). <b>(D)</b> Anti-FLAG immunoprecipitates confirm the interaction of exogenous RuvBL1 with endogenous RUVBL2. <b>(E)</b> Paraformaldehyde-fixed cells were stained with anti-FLAG antibody (green) and DAPI (blue) after 96 h of doxycycline induction. Scale bar, 5 μm. <b>(F)</b> Clonogenic survival assay after doxycycline-induced expression of the wild type or D₃₀₂N RuvBL1 variants. The experiments were performed in triplicates and a representative image is shown (quantification of the assay is shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0133576#pone.0133576.s005" target="_blank">S5 Fig</a>). <b>(G)</b> Cells were induced with doxycycline or left untreated for 96 hours. To arrest the cells in mitosis, nocodazole was added for additional 16 h. DNA content was measured by flow cytometric analysis of propidium iodide-stained cells. <b>(H)</b> Paraformaldehyde-fixed cells were stained with anti-Cyclin A antibody (red) and DAPI (blue) after 96 h of doxycycline induction and 16 h of nocodazole treatment. Scale bar, 5 μm.</p

<i>In vitro</i> phosphorylation of RUVBL1 by PLK1.

<p><b>(A)</b> Protein sequences were obtained from <a href="http://www.ncbi.nlm.nih.gov/" target="_blank">http://www.ncbi.nlm.nih.gov</a> and scanned for PLK1 consensus motifs [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0133576#pone.0133576.ref037" target="_blank">37</a>]. Corresponding sequences of human RUVBL2 are shown by comparison below the alignments. Crucial amino acids are shown in gray. <b>(B)</b> The indicated amounts of His-tagged RUVBL1 or RUVBL2 were incubated with PLK1 in the presence of [γ-³²P]ATP and protein phosphorylation was monitored by autoradiography. <b>(C)</b> Bands corresponding to phosphorylated RUVBL1 were excised and in-gel digested with trypsin. Eluted peptides were separated in two dimensions by their hydrophobicity and charge. Potential sequences were assigned to the obtained phosphopeptide patterns after autoradiography. Trypsin digestion sites are indicated with K•X. <b>(D)</b> Purified His-tagged RUVBL1 mutants were incubated with PLK1 in the presence of [γ-³²P]ATP and protein phosphorylation was monitored by autoradiography.</p

Chromosomal passenger protein-like staining of RUVBL1/2 during cell division.

<p><b>(A)</b> Methanol-fixed U2OS cells were stained with anti-RUVBL1 antibody (green) and DAPI (blue) in interphase and various stages of mitosis. At least 50 events were examined (N≥50). <b>(B)</b> Localization of GFP-mRuvBL1 at different stages of mitosis in HeLa cells. (N≥50). <b>(C)</b> Co-staining of RUVBL1 (red) and GFP-Anillin (green) in dividing cells. DNA is counterstained with DAPI (blue). Scale bar, 5 μm; (N≥50). <b>(D)</b> Co-staining of RUVBL1 (green) and β-tubulin (red) in dividing cells. DNA is counterstained with DAPI (blue). (N≥50) <b>(E)</b> Co-staining of RUVBL1 (red) and RUVBL2 (green) showing their different localization during late telophase. DNA is counterstained with DAPI (blue). (N≥50). <b>(F)</b> Whole cell extracts (1 mg) from asynchronous or double-thymidine/nocodazole-synchronized GFP-mRuvBL1 expressing HeLa cells were used for immunoprecipitation with a GFP-trap antibody. Precipitated material was analyzed with antibodies to GFP and RUVBL2. In: input (2.5%); IP: immunoprecipitated fraction; Ub: unbound fraction.</p